The purpose of this research was to improve the tumor targeting properties of radiolabeled antagonists to integrin alpha(v)beta(3) receptor by optimizing chemical parameters. In the past one year, we have focused on optimizing Tc-99m labeling methods of a second generation antagonist because Tc-99m has ideal physical properties (140 KeV, 6 hr t1/2) for scintigraphic imaging and on developing a tumor pre-targeting approach to amplify tumor-to-background signal ratio. For Tc-99m labeling, the amino terminus of 4-2-(3,4,5,6-tetrahydro-pyrimidin-2-ylamino)-ethyloxybenzoyl-2-(S)-N-(3-amino-neopenta-1-carbamyl)-aminoethylsulfonylamino--alanine hydrochloride (IAC), a carbamate derivative of a first generation antagonist (IA) was conjugated with the N-hydroxysuccinimide ester of hydrazinonicotinic acid (HYNIC) and then labeled with Tc-99m using tricine, EDDA or (tricine)(PDA) as co-ligands. Among these Tc-99m labeled IACs, Tc-99m EDDA/HYNIC-IAC produced the highest tumor uptake (2.87 +/- 0.55 %ID/g) in the receptor-positive tumor xenografted in nude mice and the lowest background radioactivity (5.03 +/- 0.58 % ID/g in intestine) in the abdomen at 2 hr post-injection; it was excreted primarily via the renal system whereas the others were excreted primarily via the hepatobiliary system, thereby producing a higher background activity in the abdomen. The nuclear imaging with Tc-99m EDDA/HYNIC-IAC clearly visualized tumors located 2 cm from the abdomen at 3 hr postinjection. For a tumor pre-targeting approach, we synthesized oligomers of a first generation antagonist (IA) using avidin (Av) as a carrier of the oligomers. The purpose was to increase the receptor-binding affinity of IA and to develop a two-step tumor pre-targeting system. The pretargeting approach involved the injection of tumor saturating dose of Av-(IA)n followed by the injection of radiolabeled biotin when the blood concentration of Av-(IA)n becomes the background concentration. The purpose was to amplify the tumor-to-background radioactivity ratios, thereby increasing imaging sensitivity and therapeutic index. Our studies demonstrated that the receptor binding affinity increased proportionally to the level of IA molecules conjugated per Av. We compared Av-(IA)n with n = <5 and >10 in vivo. The animals received ip 400 mcg of Av at -24 hr to remove endogenous biotin. We then compared pretageting parameters such as the dose and level (n= <5 and >10) of Av-(IA)n, and injection time intervals. Comparing the level (n = <5 and >10) of Av-(IA)n at 200 mcg dose injected at -4 -6 hr, n = 1015 produced higher Tumor/Blood (8.11 +/- 4.23 vs 2.3 +/- 0.93) and Tumor/Liver ratio (1.55 +/- 42 vs 0.32 +/- 0.08) than n = <5. Injection amount (>200 mcg) of Av-(IA)1015 at -6 hr and imaging at +2 hr produced the most reproducible T/B and T/L ratios. This study indicated that this pretargeting approach could amplify the signal-to-background ratio to detect tumor-induced neovasculatures.